MOLECULAR-CLONING OF HUMAN CARDIAC TROPONIN-T ISOFORMS - EXPRESSION IN DEVELOPING AND FAILING HEART

Troponin T, which links the troponin complex to tropomyosin, is found as multiple isoforms in the hearts of many animal species, Changes in isoform composition have been correlated with variation in myofilament sensitivity to calcium. In order to determine the origin of diversity of the cardiac troponin T (cTnT) isoforms indicated by existing prote in data, we have determined the sequences. and patterns of expression of mRNAs encoding troponin T in fetal and adult heart and those presen t in adult heart in end-stage failure, Three main regions of alternati ve splicing within the cTnT coding region were identified using revers e-transcriptase polymerase chain reaction (RT-PCR), Alternatively spli ced RNAs are developmentally regulated and some of the fetal forms are expressed in adult failing heart. The molecular structure of the spli ced regions was determined from cloned cDNAs and RT-PCR products. In t he 5' region of the mRNA, isoforms are generated by the inclusion or e xclusion of 15-, 3- and 27-nucleotide (nt) sequences and by the inclus ion or exclusion of a separate 3-nt sequence, In the 3' region of the mRNA, alternative splicing involves a 9-nt sequence which can be prese nt in full, in part or not at all. A further splicing site was identif ied in the central region involving a 234-nt sequence and resulting in rare but detectable mRNAs, This work demonstrates the complexity of c TnT RNA composition in human heart and provides the information necess ary to address the function of cTnT isoforms in contraction. (C) 1995 Academic Press Limited